A critical roadblock in the development of an HIV-1 vaccine is our current inability to deliver HIV-1 antigens efficiently to the immune system and to prime predictable, high frequency immune responses in humans. Recombinant Ad5 vector-based vaccines have elicited potent immune responses in preclinical studies. However, the high frequency of anti-Ad5 immunity in the developing world will likely limit the immunogenicity and clinical utility of rAd5 vaccines. We therefore propose the development of novel, replication-incompetent rAd vector-based vaccines for HIV-1. In this IPCAVD grant, we propose a focused, timeline-driven, milestone-oriented, product development program to evaluate the utility of rare serotype and molecularly engineered Ad vectors as candidate HIV-1 vaccines. We will define the optimal heterologous rAd prime-boost regimen in preclinical studies and advance this regimen into phase I and phase II clinical trials. These studies will investigate the hypothesis that these novel rAd vaccines for HIV-1 are immunogenic in the presence of anti-Ad5 immunity in both rhesus monkeys and humans. In Project 1 (Preclinical Evaluation of Novel Adenovirus Vectors), we will compare various rAd primeboost regimens in rhesus monkeys with anti-Ad5 immunity and select the optimal regimen to advance into clinical studies. We hypothesize that the optimal vector combination will involve two heterologous serotype rAd vectors that are both rare in human populations, engineered for optimal immunogenicity, derived from different Ad subfamilies, and distinct from Ad5. In Project 2 (Clinical Evaluation of Novel Adenovirus Vectors), we will conduct two phase I dose-escalation studies and a phase II heterologous rAd prime-boost study in the developing world in collaboration with the HIV Vaccine Trials Network (HVTN). Core A (Administrative Core Facility) will provide all the logistic, scientific, managerial, and financial oversight to coordinate the studies described in this IPCAVD grant. Core B (Vector Core Facility) will manufacture research-grade and clinical-grade rAd vectors and will obtain the regulatory approvals required to initiate the clinical studies. An External Steering Committee consisting of senior and distinguished leaders in the field has been assembled to supervise and to guide the overall progress of the proposed studies. By the end of the proposed 5-year period of support, we predict that we will have generated sufficient preclinical and clinical data to determine whether this vaccine strategy should be advanced into expanded clinical trials.